The present invention relates to articles made of nanostructures and, more particularly, to articles made of peptide nanostructures having sizes at least in the micrometer scale.
Material sciences involve the understanding of material characteristics as well as the development of new materials. Industrial and academic needs encourage material scientists to develop new materials having superior mechanical, electrical, optical and/or magnetic properties for many applications. Modern material sciences focus on the investigation of polymers, ceramics and semiconductors in many fluidic as well as solid forms including fibers, thin films, material bulks and the like.
Various manufacturing processes are known in the art for making synthetic fibers. Many synthetic fibers are produced by extrusion processes, in which a thick viscous liquid polymer precursor or composition is forced through one or more tiny holes of a spinneret to form continuous filaments of semi-solid polymer. As the filaments emerge from the holes of a spinneret, the liquid polymer converts first to a rubbery state which then is solidified. The process of extruding and solidifying filaments is generally known as spinning.
Wet spinning processes are typically employed with fiber-forming substances that have been dissolved in a solvent. Wet spinning techniques are preferred for spinning of high molecular weight polyamides. The spinnerets forming the filaments are submerged in a wet chemical bath, and as the filaments of the fiber-forming substances emerge from the spinnerets, they are induced to precipitate out of the solution and solidify.
In gel spinning, the polymer is not in a true liquid state during extrusion. The polymer chains are bound together at various points in liquid crystal form. This produces strong inter-chain forces in the resulting filaments that can significantly increase the tensile strength of the fibers. In addition, the liquid crystals are aligned along the fiber axis by the shear forces during extrusion. The filaments emerge with high degree of orientation relative to each other, further enhancing the strength. Typically, in gel spinning, the filaments first pass through air and then cooled in a liquid bath.
In dry spinning, the polymer is dissolved in a volatile solvent and the solution is pumped through the spinneret. As the fibers exit the spinneret, air is used to evaporate the solvent such that the fibers solidify and can be collected on a take-up wheel.
In melt spinning the polymer is melted and pumped through the spinneret. The molten fibers are cooled, solidified, and collected on a take-up wheel. Stretching of the fibers in both the molten and solid states provides for orientation of the polymer chains along the fiber axis.
Dispersion spinning is typically employed when the polymer having and infusible, insoluble and generally intractable characteristics. In this technique, the polymer is dispersed as fine particles in a chemical carrier that permit extrusion into fiber. The dispersed polymer is then caused to coalesce by a heating process and the carrier is removed by a thermal or chemical procedure.
Reaction spinning processes involve the formation of filaments from pre-polymers and monomers. The pre-polymers and monomers are further polymerized and cross-linked after the filament is formed. The reaction spinning process begins with the preparation of a viscous spinning solution, which is prepared by dissolving a low molecular weight polymer in a suitable solvent and a reactant. The spinning solution is then forced through the spinneret into a solution or being combined with a third reactant. The primary distinguishable characteristic of reaction spinning processes is that the final cross-linking between the polymer molecule chains in the filament occurs after the fibers have been spun. Post-spinning steps typically include drying and lubrication.
In tack spinning, a polymeric material in a tacky state is interposed between a foundation layer and a temporary anchorage surface. Being in a tacky state, the polymeric material adheres to the foundation layer and the temporary anchorage surface. The foundation layer is then separated from the temporary anchorage surface to produce fibers of the polymeric material. The fibers are hardened by thermal or chemical treatment, and separated from the temporary anchorage surface.
In electrospinning, a fine stream or jet of liquid is produced by pulling a small amount of charged liquefied polymer through space using electrical forces. The produced fibers are hardened and collected on a suitably located precipitation device to form a nonwoven article. In the case of a liquefied polymer which is normally solid at room temperature, the hardening procedure may be mere cooling, however other procedures such as chemical hardening or evaporation of solvent may also be employed.
Other processes for manufacturing polymeric articles include film blowing and injection molding.
In film blowing, an extruder is used to melt the polymer and pump it into a tubular die. Air blown into the center of the tube causes the melt to expand in the radial direction. The melt in thus extended in both radial and down-stream direction. The formed film is then collected by an arrangement of rollers.
In injection molding, a reciprocating or rotating screw both melts polymer pellets and provides the pressure required to inject the melt into a cold mold. The cold mold provides the article the desired shape.
In the area of thin film production, a well-known method for producing and depositing monolayers is the Langmuir-Blodgett method. In this method a monolayer of amphiphilic molecules is formed at the surface of a tank filled with a liquid sub-phase such as water. Amphiphilic molecules are those having a hydrophobic first end and a hydrophilic second end lined up side by side in a particular direction. In the Langmuir-Blodgett method, a solution of amphiphilic molecules dissolved in a solvent which is not miscible with the sub-phase liquid in the tank is spread onto the liquid surface. When the solvent evaporates, a loosely packed monolayer is formed on the surface of the sub-phase. A transition of the monolayer thus formed from a state of gas or liquid to a solid state is then achieved by compressing surface area of the layer to a predetermined surface pressure. The resulting monolayer is deposited onto the surface of a substrate by passing the substrate through the compressed layer while maintaining the layer at a predetermined surface pressure during the period of deposition.
Another method for producing a monolayer is known as self-assembling of molecules. In this method, a monolayer film is generated as a result of adsorption and bonding of suitable molecules (e.g., fatty acids, organic silicon molecules or organic phosphoric molecules) on a suitable substrate surface. The method typically involves solution deposition chemistry in the presence of water.
Over the years, extensive efforts were made to develop row materials which can be used for manufacturing fiber and films by the above techniques to provide articles having enhanced and/or application-specific characteristics. For example, one of the most studied natural fibrillar system is silk [Kaplan D L, “Fibrous proteins—silk as a model system,” Polymer degradation and stability, 59:25-32, 1998]. There are many forms of silk, of which spider silk of Nephila clavipas (the golden orb weaver) is regarded as nature's high performance fiber, with a remarkable combination of strength, flexibility, and toughness. Although assembled by non-covalent interactions, silk is stronger than steel per given fibrillar diameter but, at the same time, is much more flexible. Due to its superior mechanical properties, the spider silk can be used in many areas requiring the combination of high mechanical strength with biodegradability, e.g., in tissue engineering applications [Kubik S., “High-Performance Fibers from Spider Silk,” Angewandte Chemie International Edition, 41:2721-2723, 2002].
A known method of synthesizing spider silk material includes the introduction of a spider silk gene into a heterologous gene expression system and the secretion of spider silk protein therefrom. The protein is then processed, typically by electrospinning, to produce a fiber of enhanced mechanical properties [Jin H J, Fridrikh S V, Rutledge G C and Kaplan D L, “Electrospinning Bombyx mori silk with poly(ethylene oxide),” Biomacromolecules, 3:1233-1239, 2002].
Recently, electrospinning has been employed to fabricate virus-based composite fibers hence to mimic the spinning process of silk spiders [Lee S and Belcher A M, “Virus-Based Fabrication of Micro- and Nanofibers Using Electrospinning,” Nano letters, 4:388-390, 2004]. In this study, M13 virus was genetically modified to bind conductive and semiconductor materials, and was thereafter subjected to an electrospinning process to provide conductive and semiconductor fibers.
Other than synthesized spider silk, the electrospinning process can be applied on a diversity of polymers including polyamides, polyactides and water soluble polymer such as polyethyleneoxide [Huang Z M, Zhang Y Z, Kotaki M and Ramakrishna S., “A review on polymer nanofibers by electrospinning and their applications in nanocomposites,” Composites Science and technology, 63:2223-2253, 2003]. Heretofore, about 50 types of polymers have been successfully electrospun.
Electrospinning has also been used with carbon nanotubes to obtain super-though carbon-nanotube fibers [Dalton A B et al., “Super-tough carbon-nanotube fibres—These extraordinary composite fibres can be woven into electronic textiles,” Nature, 423:703, 2003]. By modifying a familiar method for carbon nanotubes fibers [Vigolo et al., “Macroscopic Fibers and Ribbons of Oriented Carbon Nanotubes,” Science, 17:1331-1334, 2000] the researchers were able to spin a reel of nanotube gel fiber and then convert it into 100 m length of solid nanotube composite fiber. The resulting fibers were tougher than any other known natural or synthetic organic fiber.
However, carbon nanotubes in general and carbon-nanotube fibers in particular suffer from structural deviations. Although deviations in structure can be introduced in a “controlled” manner under specific conditions, frequent uncontrollable insertion of such defects result in spatial structures with unpredictable electronic, molecular and structural properties. In addition, the production process of carbon nanotubes is very expensive and presently stands hundreds of U.S. dollars per gram.
Other known nanostructures are peptide-based nanotubular structures, made through stacking of cyclic D-, L-peptide subunits. These peptides self-assemble through hydrogen-bonding interactions into nanotubules, which in-turn self-assemble into ordered parallel arrays of nanotubes. The number of amino acids in the ring determines the inside diameter of the nanotubes obtained. Such nanotubes have been shown to form transmembrane channels capable of transporting ions and small molecules [Ghadiri, M. R. et al., Nature 366, 324-327 (1993); Ghadiri, M. R. et al., Nature 369, 301-304 (1994); Bong, D. T. et al., Angew. Chem. Int. Ed. 40:988-1011, 2001].
More recently, surfactant-like peptides that undergo spontaneous assembly to form nanotubes with a helical twist have been discovered. The monomers of these surfactant peptides, like lipids, have distinctive polar and nonpolar portions. They are composed of 7-8 residues, approximately 2 nm in length when fully extended, and dimensionally similar to phospholipids found in cell membranes. Although the sequences of these peptides are diverse, they share a common chemical property, i.e., a hydrophobic tail and a hydrophilic head. These peptide nanotubes, like carbon and lipid nanotubes, also have a very high surface area to weight ratio. Molecular modeling of the peptide nanotubes suggests a possible structural organization [Vauthey (2002) Proc. Natl. Acad. Sci. USA 99:5355; Zhang (2002) Curr. Opin. Chem. Biol. 6:865]. Based on observation and calculation, it is proposed that the cylindrical subunits are formed from surfactant peptides that self-assemble into bilayers, where hydrophilic head groups remain exposed to the aqueous medium. Finally, the tubular arrays undergo self-assembly through non-covalent interactions that are widely found in surfactant and micelle structures and formation processes.
Peptide based bis(N-α-amido-glycyglycine)-1,7-heptane dicarboxylate molecules were also shown to be assembled into tubular structures [Matsui (2000) J. Phys. Chem. B 104:3383].
When the crystal structure of di-phenylalanine peptides was determined, it was noted that hollow nanometric channels are formed within the framework of the macroscopic crystal [Gorbitz et al., Chemistry 7(23):5153-9, 2001]. However, no individual nanotubes could be formed by crystallization, as the crystallization conditions used in this study included evaporation of an aqueous solution at 80° C. No formation of discrete nano-structures was reported under these conditions.
International Patent Application Nos. IL03/01045 and IL2004/000012 (see also Reches M and Gazit E, “Casting metal nanowires within discrete self-assembled peptide nanotubes,” Science, 300:625-627, 2003), disclose a new procedure for making peptide nanostructures that show many ultrastructural and physical similarities to carbon nanotubes. These peptide nanostructures are self assembled by diphenylalanine, the core recognition motif of the β-amyloid peptide [Findeis et al., “Peptide inhibitors of beta amyloid aggregation,” Biochemistry, 38:6791, 1999; Tjernberg et al., “Arrest of -Amyloid Fibril Formation by a Pentapeptide Ligand,” J. Biol. Chem., 271:8545-8548, 1996; and Soto et al., “Beta-sheet breaker peptides inhibit fibrillogenesis in a rat brain model of amyloidosis: implications for Alzheimer's therapy,” Nature Medicine, 4:822-826, 1998].
The self-assembled peptide nanostructures are well ordered assemblies of various shapes with persistence length on the order of micrometers. The formation of the peptide nanostructures is very efficient and the nanostructures solution is very homogeneous. Similar to carbon nanotubes, the peptide nanostructures are formed as individual entities. For industrial applications, the self-assembled peptide nanostructures are favored over carbon nanotubes and spider silk from standpoint of cost, production means and availability.
There is thus a widely recognized need for, and it would be highly advantageous to have macroscopic and microscopic articles exploiting the advantages of self-assembled peptide nanostructures.